Current policy to protect Americans against smallpox mandates immunization of naive and previously vaccinated adults with live vaccinla virus (VV). Knowledge of adaptive immunity to VV is limited to descriptions of neutralizing antibodies, cytotoxic T lymphocyte (CTL) activity, and lymphocyte proliferation responses, largely because widespread vaccination ceased before the current era of molecular immunology. Research in this project will examine the evolution of VV-specific memory T cells in newly vaccinated and re-vaccinated (boosted) adults. The relationship of VV-specific adaptive immunity to viral load and innate immunity will be determined in conjunction with projects by Storch and Stanley. The specific aims are to: 1. Identify immunodominant VV epitopes that induce CD8+ T cell memory. Artificial neural networks and computational algorithms will initially be used to select peptide epitopes, followed by evaluation of peptide binding to HLA class I alleles and assessment of the ability of peptides to stimulate IFN-gamma and IL-2 responses by HLA-matched and HLA-mismatched primary vaccinees and adults given booster doses of VV. 2. Determine the functional phenotype of CD8+ cells during the inductive and memory phases of adaptive immunity to VV. HLA class I tetramer-peptide complexes will be used to identify and quantify CD8+ cells bearing TCR specific for VV epitopes before vaccination and 9 days, 6, 12, and 24 months post-vaccination. Lymphocytes producing putative mediators of immunity (IFN-gamma, perforin, MIP-1alpha) and bearing memory markers (CD45RO, CD45RA, CCR7) will be evaluated prospectively in primary vaccinees and boosted adults. 3. Evaluate the clonotypic repertoire of VV-specific CD8+ cells following primary vaccination and boosting of previously vaccinated adults. TCR Vbeta usage and CDR3 length and sequence polymorphism will be evaluated prospectively in selected HLA-A2+ and HLA-A11+ individuals.